Devices and methods for detecting light have been known in practice for many years, and are used in laser-scanning microscopes, for example. In this case, corresponding devices that detect the detection signal from the sample to be microscopically examined are of key importance to the image quality. This primarily applies to comparatively weak detection signals, as are typical in (confocal) fluorescence microscopy, SHG microscopy or Raman microscopy.
Two parameters are of particular importance for light detectors, namely the detector noise and the quantum yield, i.e. the detection efficiency. Here, the quantum yield describes the proportion of light incident on the detector that actually generates a usable electrical signal. The noise describes the electronic base signal that is superimposed on the actual detection signal in an interfering manner. The ratio between these two values, known as the signal to noise ratio (SNR), is one of the key parameters of a light detector.
In practice, photomultipliers (PMTs) have been the dominant light detectors in laser-scanning microscopy for many years. In comparison with semiconductor-based detectors, e.g. photodiodes, PMTs have a lower quantum yield. Due to the low noise thereof, they provide a very good SNR, however. Furthermore, improved variants having a GaAsP (gallium arsenide phosphide) layer as a light-sensitive medium have become available in recent years.
Furthermore, it has been known for a number of years to use semiconductor detectors in fluorescence microscopy as an alternative. Here, “single-photon avalanche diodes” (SPAD) are of particular significance. The SPADs operate in Geiger mode. In this case, a cutoff voltage is applied to the SPADs, which is slightly above the breakdown voltage. The breakdown voltage is several hundred volts in this case.
In this mode, an absorbed photon generates an electron-hole pair in the semiconductor, which pair is accelerated by the strong electrical field and carries out further impact ionizations. This process continues in the manner of an avalanche and triggers a measurable charge avalanche that is amplified by a factor of several million. Therefore, individual absorbed photons can be measured, meaning that these detectors can be used for measuring extremely low amounts of light, as are common in fluorescence microscopy, for example.
In this process, a single photon leads to an electrical discharge, which is measured in the form of a short voltage pulse. Here, there are essentially two measurement modes. In the digital measurement mode, the voltage pulses are counted, the rising voltage edge being used as a triggering counter signal. Alternatively, in what is known as the analog measurement mode, the charge can be integrated by means of a measuring resistor and the accumulated charge of all the pulses is used as a measurement signal. Typically, the integrated charge of all the pulses in a set time period (pixel exposure time) is then digitalized by an analog/digital converter for further digital processing.
Irrespective of the selected measurement mode, one problem with SPADs is that signal saturation occurs. Specifically, the measurement signal no longer rises to the same extent when the amount of light incident on the detector increases. The desired linear relationship between the input signal and the output signal thus no longer exists. The saturation occurs because, during an avalanche discharge of the SPADs, another absorbed photon cannot trigger a simultaneous, second avalanche. Therefore, after pulse triggering, there is some dead time of the SPADs during which detection cannot take place. This dead time corresponds to the time that is required to recharge the charge carrier in the semiconductor that has been depleted during the avalanche discharge. Large amounts of light in which a plurality of photons are incident within the dead time therefore can no longer be completely detected and the detector displays a non-linear saturation characteristic curve.
Since the low dynamic range of the detectors resulting therefrom having a maximum count rate of some 106 to 107 photons per second is a problem in these highly sensitive detectors, what are known as SPAD arrays have been developed in recent years. These are available for example from the manufacturer Hamamatsu Photonics K.K. under the name multi-pixel photon counting detector, or MPPC detector. In the literature, these detectors are, inter alia, also referred to as silicon photomultipliers (SiPM). The function of corresponding detectors is for example described at
<<https://www.hamamatsu.com/resources/pdf/ssd/mppc_techinfo_e.pdf>>.
The basic principle of an SPAD array involves a plurality of individual SPADs being connected in parallel to form a field. If a photon is incident on an individual SPAD, then, due to its dead time, it is no longer sensitive for a period of typically a few nanoseconds. Other SPADs on which another photon is incident within this period or even simultaneously can however detect said photon and generate a measurable charge pulse. Therefore, a pulse sequence having a higher count rate than in an individual SPAD can be produced at the detector output.
Therefore, it is known from the prior art to distribute the entirety of the detection light across a plurality of SPADs connected in parallel. This is advantageous in that only a fraction of the detection light impinges on individual SPADs and these SPADs are therefore saturated later. Furthermore, additional SPADs that are ready to receive are available during the dead time of an individual SPAD. The dynamic range of these detectors is therefore significantly increased, depending on the number of SPADs that are connected in parallel. Commercially available SPAD arrays have 20×20 or more SPADs, for example.
However, the known SPAD arrays are also problematic in that they exhibit saturation behavior. Saturation may occur if too many photons are incident on the same SPAD within the dead time. In this case, the saturation is similar to the saturation of a single SPAD. Furthermore, saturation may also occur in the digital detection mode if the rising trigger edges of a pulse do not lead to the digital counter being triggered again during a previous pulse, since the voltage level thereof is above the voltage threshold for triggering the counter (known as the trigger level), and therefore is not detected as a pulse edge by the counting unit.
The two above-mentioned effects also lead to saturation of SPAD arrays. It is known in practice that, for example from approx. 108 incident photons per second, the output signal (the number of electrical discharges) assumes an approximately constant value, such that it is no longer possible to precisely measure the amount of light. Provided that the characteristic curve, which is specific to a given detector design, is known, it can be linearized by computational correction. In the range of almost complete saturation (for example slightly above 1011photons per second), computational correction is no longer possible in a sufficiently precise manner, however. Since a characteristic curve in the analog measurement mode displays an almost identical curve as in the digital measurement mode, a distinction is not made in the following between the different causes of saturation.
Although connecting many SPADs in parallel to form an SPAD array therefore constitutes an improvement on the problem of saturation, this problem still persists. Therefore, the dynamic range of known SPAD arrays is still further below the dynamic range of photomultipliers, which are therefore still used for detecting low amounts of light despite the detection efficiency thereof being worse than that of SPAD detectors.